RGD Peptide-Conjugated Selenium Nanocomposite Inhibits Human Glioma Growth by Triggering Mitochondrial Dysfunction and ROS-Dependent MAPKs Activation

Chemotherapy is still one of the most common ways to treat human glioblastoma in clinic. However, severe side effects limited its clinic application. Design of cancer-targeted drugs with high efficiency and low side effect is urgently needed. Herein, silver nanoparticles (Ag NPs) and nano-selenium (Se NPs) conjugated with RGD peptides (Ag@Se@RGD NPs) to target integrin high-expressed glioma were designed. The results found that Ag@Se@RGD NPs displayed stable particle size and morphology in physiological condition, and induced significant integrin-targeted intracellular uptake. Ag@Se@RGD NPs in vitro dose-dependently inhibited U251 human glioma cells growth by induction of cells apoptosis through triggering the loss of mitochondrial membrane potential, overproduction of reactive oxygen species (ROS), and MAPKs activation. However, ROS inhibition dramatically attenuated Ag@Se@RGD NPs-induced MAPKs activation, indicating the significant role of ROS as an early apoptotic event. Importantly, Ag@Se@RGD NPs administration in vivov effectively inhibited U251 tumor xenografts growth by induction of apoptosis through regulation MAPKs activation. Taken together, our findings validated the rational design that Ag-Se NPs conjugated with RGD peptides was a promising strategy to combat human glioma by induction of apoptosis through triggering mitochondrial dysfunction and ROS-dependent MAPKs activation.

Exploiting maleimide-functionalized hyaluronan hydrogels to test cellular responses to physical and biochemical stimuli

Current in vitro 3D models of liver tissue have been limited by the inability to study the effects of specific extracellular matrix (ECM) components on cell phenotypes. This is in part due to limitations in the availability of chemical modifications appropriate for this purpose. For example, hyaluronic acid (HA), which is a natural ECM component within the liver, lacks key ECM motifs (e.g., RGD peptides) that support cell adhesion. However, the addition of maleimide (Mal) groups to HA could facilitate the conjugation of ECM biomimetic peptides with thiol-containing end groups. In this study, we characterized a new crosslinkable hydrogel (i.e., HA-Mal) that yielded a simplified ECM-mimicking microenvironment supportive of 3D liver cell culture. We then performed a series of experiments to assess the impact of physical and biochemical signaling in the form of RGD peptide incorporation and TGF- ß supplementation, respectively, on hepatic functionality. Hepatic stellate cells (i.e., LX-2) exhibited increased cell-matrix interactions in the form of cell spreading and elongation within HA-Mal matrices containing RGD peptides, enabling physical adhesions, whereas hepatocyte-like cells (HepG2) had reduced albumin and urea production. We further exposed the encapsulated cells to soluble TGF-ß to elicit a fibrosis-like state. In the presence of TGF-ß biochemical signals, LX-2 cells became activated and HepG2 functionality significantly decreased in both RGD-containing and RGD-free hydrogels. Altogether, in this study we have developed a hydrogel biomaterial platform that allows for discrete manipulation of specific ECM motifs within the hydrogel to better understand the roles of cell-matrix interactions on cell phenotype and overall liver functionality.

Non-Cationic RGD-Containing Protein Nanocarrier for Tumor-Targeted siRNA Delivery

Despite the recent successes in siRNA therapeutics, targeted delivery beyond the liver remains the major hurdle for the widespread application of siRNA in vivo. Current cationic liposome or polymer-based delivery agents are restricted to the liver and suffer from off-target effects, poor clearance, low serum stability, and high toxicity. In this study, we genetically engineered a non-cationic non-viral tumor-targeted universal siRNA nanocarrier (MW 26 KDa). This protein nanocarrier consists of three function domains: a dsRNA binding domain (dsRBD) (from human protein kinase R) for any siRNA binding, 18-histidine for endosome escape, and two RGD peptides at the N- and C-termini for targeting tumor and tumor neovasculature. We showed that cloned dual-RGD-dsRBD-18his (dual-RGD) protein protects siRNA against RNases, induces effective siRNA endosomal escape, specifically targets integrin αvβ3 expressing cells in vitro, and homes siRNA to tumors in vivo. The delivered siRNA leads to target gene knockdown in the cell lines and tumor xenografts with low toxicity. This multifunctional and biomimetic siRNA carrier is biodegradable, has low toxicity, is suitable for mass production by fermentation, and is serum stable, holding great potential to provide a widely applicable siRNA carrier for tumor-targeted siRNA delivery.

Function and Mechanism of RGD in Bone and Cartilage Tissue Engineering

Bone and cartilage injury is common, tissue engineered scaffolds are potential means to repair. Because most of the scaffold materials used in bone and cartilage tissue engineering are bio-inert, it is necessary to increase the cellular adhesion ability of during tissue engineering reconstruction. The Arginine - Glycine - Aspartic acid (Arg-Gly-Asp, RGD) peptide family is considered as a specific recognition site for the integrin receptors. Integrin receptors are key regulators of cell-cell and cell-extracellular microenvironment communication. Therefore, the RGD polypeptide families are considered as suitable candidates for treatment of a variety of diseases and for the regeneration of various tissues and organs. Many scaffold material for tissue engineering and has been approved by US Food and Drug Administration (FDA) for human using. The application of RGD peptides in bone and cartilage tissue engineering was reported seldom. Only a few reviews have summarized the applications of RGD peptide with alloy, bone cements, and PCL in bone tissue engineering. Herein, we summarize the application progress of RGD in bone and cartilage tissue engineering, discuss the effects of structure, sequence, concentration, mechanical stimulation, physicochemical stimulation, and time stimulation of RGD peptide on cells differentiation, and introduce the mechanism of RGD peptide through integrin in the field of bone and cartilage tissue engineering.

Cyclic Peptide Modified Gold Clusters Induce Lung Tumor Cell Apoptosis via Generating Intracellular Oxidative Stress

Metastatic lung cancer is the leading cause of death for cancer patients. Although many chemical drugs were developed for cancer treatment, metastatic cancer mortality did not decrease significantly. In this article, we designed an Au clusters (AuCs) modified by cyclic RGD peptides which well target the integrin of human lung carcinoma cells (A549). The RGD-AuCs could well induce A549 cells apoptosis, but have no cytotoxicity on the human bronchial epithelial cells (16HBE), which are normal cells support respiratory system. The AuCs could be internalized and localized in the lysosomes of A549 tumor cells and further release into cytoplasma. We found the ROS level was increased by AuCs, and such high ROS level finally leads to depolarization of mitochondria. Eventually, the AuCs stimulating mitochondria related apoptosis pathway to induce A549 tumor cells apoptosis. We deduce the gold clusters would be an effective therapeutic candidate to against metastatic lung tumor in the future studies.

There is a world beyond αvβ3-integrin: Multimeric ligands for imaging of the integrin subtypes αvβ6, αvβ8, αvβ3, and α5β1 by positron emission tomography

Background In the context of nuclear medicine and theranostics, integrin-related research and development was, for most of the time, focused predominantly on 'RGD peptides' and the subtype αvβ3-integrin. However, there are no less than 24 known integrins, and peptides without the RGD sequence as well as non-peptidic ligands play an equally important role as selective integrin ligands. On the other hand, multimerization is a well-established method to increase the avidity of binding structures, but multimeric radiopharmaceuticals have not made their way into clinics yet. In this review, we describe how these aspects have been interwoven in the framework of the German Research Foundation's multi-group interdisciplinary funding scheme CRC 824, yielding a series of potent PET imaging agents for selective imaging of various integrin subtypes. Results The gallium-68 chelator TRAP was utilized to elaborate symmetrical trimers of various peptidic and non-peptidic integrin ligands. Preclinical data suggested a high potential of the resulting Ga-68-tracers for PET-imaging of the integrins α5β1, αvβ8, αvβ6, and αvβ3. For the first three, we provide some additional immunohistochemistry data in human cancers, which suggest several future clinical applications. Finally, application of αvβ3- and αvβ6-integrin tracers in pancreatic carcinoma patients revealed that unlike αvβ3-targeted PET, αvβ6-integrin PET is not characterized by off-target uptake and thus, enables a substantially improved imaging of this type of cancer. Conclusions Novel radiopharmaceuticals targeting a number of different integrins, above all, αvβ6, have proven their clinical potential and will play an increasingly important role in future theranostics.

Synthesis and Evaluation of a Dimeric RGD Peptide as a Preliminary Study for Radiotheranostics with Radiohalogens

We recently developed 125I- and 211At-labeled monomer RGD peptides using a novel radiolabeling method. Both labeled peptides showed high accumulation in the tumor and exhibited similar biodistribution, demonstrating their usefulness for radiotheranostics. This study applied the labeling method to a dimer RGD peptide with the aim of gaining higher accumulation in tumor tissues based on improved affinity with αvβ3 integrin. We synthesized an iodine-introduced dimer RGD peptide, E[c(RGDfK)] (6), and an 125/131I-labeled dimer RGD peptide, E[c(RGDfK)]{[125/131I]c[RGDf(4-I)K]} ([125/131I]6), and evaluated them as a preliminary step to the synthesis of an 211At-labeled dimer RGD peptide. The affinity of 6 for αvβ3 integrin was higher than that of a monomer RGD peptide. In the biodistribution experiment at 4 h postinjection, the accumulation of [125I]6 (4.12 ± 0.42% ID/g) in the tumor was significantly increased compared with that of 125I-labeled monomer RGD peptide (2.93 ± 0.08% ID/g). Moreover, the accumulation of [125I]6 in the tumor was greatly inhibited by co-injection of an excess RGD peptide. However, a single injection of [131I]6 (11.1 MBq) did not inhibit tumor growth in tumor-bearing mice. We expect that the labeling method for targeted alpha therapy with 211At using a dimer RGD peptide could prove useful in future clinical applications.

Synthetic Integrin-Targeting Dextran-Fc Hybrids Efficiently Inhibit Tumor Proliferation In Vitro

Herein, we present the design, synthesis, and biological evaluation of novel integrin-targeting molecular hybrids combining RGD peptides and a potent cytotoxin presented on dextran polysaccharides. Based on an aglycosylated Fc as a centerpiece, endosomal-cleavable cytotoxic agent monomethyl auristatin E (MMAE) and dextran as multimerization site were covalently connected by two bioorthogonal enzyme-mediated reactions site-specifically. Decoration of dextran with cyclic RGD peptides, introduced by copper “click” reaction, resulted in the final constructs with the potential to kill integrin-overexpressing tumor cells. We found that these modifications had little impact on the stability of the Fc scaffold and the RGD-bearing construct showed good binding properties of αvβ3-expressing U87MG cells. Furthermore, the construct showed a remarkable antiproliferative activity. These results demonstrate the general capability of our design to provoke receptor-mediated endocytosis upon binding to the cellular surface, followed by endosomal cleavage of the linkage between Fc-dextran and MMAE and its subsequent release. Our approach opens new avenues to transcribe small molecule binders into tailor-made multimeric molecular hybrids with antitumor potential.